A widely distributed diheme enzyme from Burkholderia that displays an atypically stable bis-Fe(IV) state

Bacterial diheme peroxidases represent a diverse enzyme family with functions that range from hydrogen peroxide (H 2 O 2 ) reduction to post-translational modifications. By implementing a sequence similarity network (SSN) of the bCCP_MauG superfamily, we present the discovery of a unique diheme peroxidase BthA conserved in all Burkholderia . Using a combination of magnetic resonance, near-IR and Mössbauer spectroscopies and electrochemical methods, we report that BthA is capable of generating a bis- Fe(IV) species previously thought to be a unique feature of the diheme enzyme MauG. However, BthA is not MauG-like in that it catalytically converts H 2 O 2 to water, and a 1.54-Å resolution crystal structure reveals striking differences between BthA and other superfamily members, including the essential residues for both bis- Fe(IV) formation and H 2 O 2 turnover. Taken together, we find that BthA represents a previously undiscovered class of diheme enzymes, one that stabilizes a bis- Fe(IV) state and catalyzes H 2 O 2 turnover in a mechanistically distinct manner. The diheme enzyme MauG forms a bis -Fe(IV) state. Here the authors identify and determine the structure of BthA, a diheme peroxidase conserved in all Burkholderia and show that BthA also forms a bis -Fe(IV) species but mechanistically differs from MauG by combining magnetic resonance, near-IR and Mössbauer spectroscopies and electrochemical methods.